Efficient gene disruption in Saccharomyces cerevisiae using marker cassettes with long homologous arms prepared by the restriction-free cloning strategy

研究领域[WOS]:

Biotechnology & Applied Microbiology

英文摘要:

Here we report an improved method for targeted gene disruption with high efficiency in S. cerevisiae, where the selection markers with long homologous arms are defined by the choice of the primer binding sites at the target locus and the disruption cassettes are constructed by restriction-free (RF) cloning strategy. Three genes, SAM1, IDH1 and IDH2, were disrupted with this method and the disruption efficiencies of SAM1 was improved several folds with much lower false-positive rates compared to the conventional one-step PCR-based gene disruption method. This approach for gene disruption cassettes construction with long flanking homologous arms may be readily applicable to facilitate targeted gene disruption in other non-conventional yeasts and fungi.